FRDA is an inherited mitochondrial disease resulting from a triplet nucleotide expansion in the first intron of the gene for frataxin, a protein involve in iron sulfur cluster (ISC) formation. Patients with FRDA have decreased levels of frataxin and secondarily ISC-containing proteins, resulting in the degeneration of dorsal root ganglion cells, the dentate nucleus of the cerebellum, selected other neurons, cardiomyocytes, and -islet cells. Clinically, these changes manifest in a characteristic ataxia, dysarthria, hypertrophic cardiomyopathy and diabetes mellitus. Although increased reactive oxygen species (ROS) formation and oxidative stress have been proposed as pathogenic mechanisms of FRDA based on results from cellular models, treatments aimed at these have been minimally successful. In addition, as frataxin is a ubiquitously expressed protein, the mechanism behind the exquisite tissue selectivity in FRDA is unexplained. An alternative hypothesis to ROS production for the pathophysiology of FRDA is that insufficient bioenergetic capacity with downstream metabolic dysfunction plays a crucial role in the disease. In addition to their role the in the electron transport chain, iron-sulfur containing enzymes are also involved in the Krebs cycle and fatty acid breakdown. Consequently, the metabolic functions of mitochondria might be crucially compromised in FRDA, leading to tissue selective metabolic dysfunction. In order initially to define such abnormalities, we have used rigorous stable isotope dilution and isotopic tracer liquid chromatography-mass spectrometry techniques to examine metabolism in FRDA. Using Stable Isotope Labeling by Essential nutrients in Cell culture (SILEC), a novel stable isotope dilution LC-MS method for assessing metabolic changes reflected revealed by intracellular levels of short chain acyl-coenzyme (CoA) thioesters, we have found that decreased frataxin expression results in changes of intracellular levels of specific CoA thioesters. In addition, we have identified a decreased metabolism through fatty acid pathways from palmitate to Acetyl CoA in platelets and fibroblasts. Such observations may be useful as biomarkers of disease activity, and potentially for developing therapeutic approaches. In this proposal, we will expand the understanding fatty acid metabolism in FRDA by examining other metabolism other substrates, in different cellular models, and with a broader scope of FRDA patients. We will examine metabolism of other substrates, as well as the tissue specificity of metabolic changes. Such measurements will develop biomarkers that can be used in clinical studies, including therapeutic trials, in FRDA. This will be tested in the final aim. Overall this approach will have immediate impact on ongoing trials in FRDA and potentially revise current pathophysiological concepts of the disorder and related diseases.